Malfunctioning lymphatics and lymphatic function are strongly implicated in metastatic cancer, asthma, diabetes, and obesity. Yet despite the importance of the lymphatics in these diseases, diagnostic imaging approaches to non-invasively monitor lymph function are non-existent. Recently, we reported the ability to non-invasively and dynamically image lymph function in the intact swine as well as in humans using a non- specific near-infrared fluorophore, IC-Green, as well as a molecular stain of lymph endothelium that targets the lymph vascular endothelium receptor, LYVE-1. In addition, upon developing imaging agents that molecularly target activated lymph endothelium, we hypothesize that we can image lymphangiogenesis. Our results demonstrate the feasibility of monitoring lymph flow and its molecular markers for the diagnosis and management of lymph diseases in a population of patients who suffer from primary and secondary lymphedema. In this research application, we hypothesize that: near infrared (NIR) optical methods can be used to image lymph flow, function, and remodeling to provide diagnostics and information on response to therapy. Specifically, we seek to: (1) Refine imaging hardware and develop software methodologies for automated imaging and analysis of lymph function in preclinical and clinical imaging studies; (2) Develop, synthesize, and validate molecularly targeting imaging agents that are specific to LYVE-1, hyaluronidase, and alpha-9 integrins that are involved in the process of lymphedema and lymphangiogenesis; (3) Develop functional lymph imaging methodologies in an immunocompetent SKH-1 mouse model of acquired lymphedema in order to assess the temporal progression of lymphedema using trace amounts of non-specific IC-Green and IRDye800CW to assess lymphatic function, and then to use molecular imaging agents that target LYVE-1, hyaluronan degradation, and alpha-9, alpha-v integrins expressed on proliferating LEC and BEC proliferation to track the progression and response to therapy; and finally to (4) Conduct imaging on human subject volunteers to establish baseline lymph function imaging parameters as well as on lymphedema patients to assess alteration in lymph function imaging parameters with disease progression and response/non-response to conventional therapy. The proposed work utilizes new developmental instrumentation and imaging agents, established mouse models of lymphedema, and a clinical trial team to translate discoveries in lymph imaging to patients. Project Narrative: Malfunctioning lymphatics and lymphatic function are strongly implicated in metastatic cancer, asthma, diabetes, and obesity. Yet despite the importance of the lymphatics in these diseases, diagnostic imaging approaches to non-invasively monitor lymph function are non-existent. In this work we develop new imaging techniques to assess lymph function and molecular markers in lymph disease and health.